We are interested in understanding the induction and characterization of anti-DNA antibodies in systemic lupus erythematosus (SLE). Of particular interest is the role of foreign antigen in triggering the generation of pathogenic autoreactive B-cells during the somatic diversification of the antibody response. Since we have shown that anti-PC antibodies can mutate to dsDNA binding in vitro and in vivo, we will immunize with PC-carrier the T15i transgenic mouse developed by Rajewsky and colleagues, in which the rearranged S107 T15 heavy chain V region is inserted by homologous recombination into Ig heavy chain locus upstream of the mu constant region. We propose to study how frequently somatic mutation of the T15 heavy chain generates B-cells making antibodies that bind dsDNA during the response of nonautoimmune and autoimmune mice to PC. We will generate hybridomas with NSObcl-2, a new fusion partner that rescues B-cells programmed for apoptosis. We will also immunize non-autoimmune and autoimmune mice with TNP and phenylarsonate to see if foreign antigens routinely activate anti-self antibodies, as the B-cell response to these haptens has been demonstrated to be clonally related to an anti-DNA response. We will examine the spectrum of amino acid changes that can lead to dsDNA binding in the absence of the selective forces that exist in vivo by generating DNA binding antibodies from the same S107 V1 heavy chain gene in a new tissue culture system where V region hypermutation occurs. These studies should reveal the role of environmental antigens and somatic mutation in the generation of anti-dsDNA antibodies in vivo and provide insight into the mechanisms of peripheral tolerance that regulate the expression of such antibodies in nonautoimmune mice as well as the potential defects in this regulation in autoimmune mice.